Photobiomodulation of mineralisation in mesenchymal stem cells.

Mesenchymal stem cells (MSCs) and photobiomodulation (PBM) both offer significant therapeutic potential in regenerative medicine. MSCs have the ability to self-renew and differentiate; giving rise to multiple cellular and tissue lineages that are utilised in repair and regeneration of damaged tissues. PBM utilises light energy delivered at a range of wavelengths to promote wound healing. The positive effects of light on MSC proliferation are well documented; and recently, several studies have determined the outcomes of PBM on mineralised tissue differentiation in MSC populations. As PBM effects are biphasic, it is important to understand the underlying cellular regulatory mechanisms, as well as, provide accurate details of the irradiation conditions, to optimise and standardise outcomes. This review article focuses on the use of red, near-infra-red (R/NIR) and blue wavelengths to promote the mineralisation potential of MSCs; and also reports on the possible molecular mechanisms which underpin transduction of these effects. A variety of potential photon absorbers have been identified which are reported to mediate the signalling mechanisms, including respiratory chain enzymes, flavins, and cryptochromes. Studies report that R/NIR and blue light stimulate MSC differentiation by enhancing respiratory chain activity and increasing reactive oxygen species levels; however, currently, there are considerable variations between irradiation parameters reported. We conclude that due to its non-invasive properties, PBM may, following optimisation, provide an efficient therapeutic approach to clinically support MSC-mediated hard tissue repair. However, to optimise application, further studies are required to identify appropriate light delivery parameters, as well as elucidate the photo-signalling mechanisms involved.

Potential for direct application of blue light for photo-disinfection of dentine.

The direct application of light for photo-disinfection potentially provides a safe and novel modality to inhibit or eliminate cariogenic bacteria residing upon and within dentine. This study aimed to both; characterize the pattern of transmission of 405 nm light through molar dentine at different tooth locations, as well as, determine the irradiation parameters that are antibacterial for Streptococcus mutans under various growth conditions, including lawns, planktonic cultures, and biofilms. To determine the amount of light (405 nm) transmitted at different anatomical tooth locations; irradiance values were recorded after blue light (470-4054 mW/cm2) had traversed through occlusal, oblique, and buccal dentine sections; and three thicknesses - 1, 2 and 3 mm were investigated. To determine tubular density; scanning electron micrographs from 2 mm outer (dentine-enamel junction) and inner (pulp) dentine sections were analysed. For photo-disinfection studies; S. mutans was irradiated using the same 405 nm wavelength light at a range of doses (110-1254 J/cm2) in both biofilm and planktonic cultures. The inhibitory effect of the irradiation on bacterial lawns was compared by measuring zones of inhibition; and for planktonic cultures both spectrophotometric and colony forming unit (CFU) assays were performed. A live/dead staining assay was utilised to determine the effect of irradiation on bacterial viability in mature biofilms. Data indicated that increasing dentine thickness decreased light transmission significantly irrespective of its orientation. Occlusal and oblique samples exhibited higher transmission compared with buccal dentine. Oblique dentine 405 nm light transmission was comparable with that of occlusal dentine independent of section thickness. An increased tubule density directly positively correlated with light transmission. Irradiation at 405 nm inhibited S. mutans growth in both biofilm and planktonic cultures and a dose response relationship was observed. Irradiation at doses of 340 and 831 J/cm2 led to significant reductions in bacterial growth and viability; as determined by CFU counting and live/dead staining. Data suggests that phototherapy approaches utilising a 405 nm wavelength have therapeutic potential to limit cariogenic bacterial infections both at the surface and within dentine.

Development and application of LED arrays for use in phototherapy research.

Lasers/LEDs demonstrate therapeutic effects for a range of biomedical applications. However, a consensus on effective light irradiation parameters and efficient and reliable measurement techniques remain limited. The objective here is to develop, characterise and demonstrate the application of LED arrays in order to progress and improve the effectiveness and accuracy of in vitro photobiomodulation studies. 96-well plate format LED arrays (400-850 nm) were developed and characterised to accurately assess irradiance delivery to cell cultures. Human dental pulp cells (DPCs) were irradiated (3.5-142 mW/cm2 : 15-120 s) and the biological responses were assessed using MTT assays. Array calibration was confirmed using a range of optical and analytical techniques. Multivariate analysis of variance revealed biological responses were dependent on wavelength, exposure time and the post-exposure assay time (P < 0.05). Increased MTT asbsorbance was measured 24 h post-irradiation for 30 s exposures of 3.5 mW/cm2 at 470, 527, 631, 655, 680, 777, 798 and 826 nm with distinct peaks at 631 nm and 798 nm (P < 0.05). Similar wavelengths were also effective at higher irradiances (48-142 mW/cm2 ). LED arrays and high throughput assays provide a robust and reliable platform to rapidly identify irradiation parameters which is both time- and cost-effective. These arrrays are applicable in photobiomodulation, photodynamic therapy and other photobiomedical research.

The dark art of light measurement: accurate radiometry for low-level light therapy.

Lasers and light-emitting diodes are used for a range of biomedical applications with many studies reporting their beneficial effects. However, three main concerns exist regarding much of the low-level light therapy (LLLT) or photobiomodulation literature; (1) incomplete, inaccurate and unverified irradiation parameters, (2) miscalculation of 'dose,' and (3) the misuse of appropriate light property terminology. The aim of this systematic review was to assess where, and to what extent, these inadequacies exist and to provide an overview of 'best practice' in light measurement methods and importance of correct light measurement. A review of recent relevant literature was performed in PubMed using the terms LLLT and photobiomodulation (March 2014-March 2015) to investigate the contemporary information available in LLLT and photobiomodulation literature in terms of reporting light properties and irradiation parameters. A total of 74 articles formed the basis of this systematic review. Although most articles reported beneficial effects following LLLT, the majority contained no information in terms of how light was measured (73%) and relied on manufacturer-stated values. For all papers reviewed, missing information for specific light parameters included wavelength (3%), light source type (8%), power (41%), pulse frequency (52%), beam area (40%), irradiance (43%), exposure time (16%), radiant energy (74%) and fluence (16%). Frequent use of incorrect terminology was also observed within the reviewed literature. A poor understanding of photophysics is evident as a significant number of papers neglected to report or misreported important radiometric data. These errors affect repeatability and reliability of studies shared between scientists, manufacturers and clinicians and could degrade efficacy of patient treatments. Researchers need a physicist or appropriately skilled engineer on the team, and manuscript reviewers should reject papers that do not report beam measurement methods and all ten key parameters: wavelength, power, irradiation time, beam area (at the skin or culture surface; this is not necessarily the same size as the aperture), radiant energy, radiant exposure, pulse parameters, number of treatments, interval between treatments and anatomical location. Inclusion of these parameters will improve the information available to compare and contrast study outcomes and improve repeatability, reliability of studies.

Developments in low level light therapy (LLLT) for dentistry.

OBJECTIVES: Low level light/laser therapy (LLLT) is the direct application of light to stimulate cell responses (photobiomodulation) in order to promote tissue healing, reduce inflammation and induce analgesia. There have been significant studies demonstrating its application and efficacy at many sites within the body and for treatment of a range of musculoskeletal injuries, degenerative diseases and dysfunction, however, its use on oral tissues has, to date, been limited. The purpose of this review is to consider the potential for LLLT in dental and oral applications by providing background information on its mechanism of action and delivery parameters and by drawing parallels with its treatment use in analogous cells and tissues from other sites of the body. METHODS: A literature search on Medline was performed on laser and light treatments in a range of dental/orofacial applications from 2010 to March 2013. The search results were filtered for LLLT relevance. The clinical papers were then arranged to eight broad dental/orofacial categories and reviewed. RESULTS: The initial search returned 2778 results, when filtered this was reduced to 153. 41 were review papers or editorials, 65 clinical and 47 laboratory studies. Of all the publications, 130 reported a positive effect in terms of pain relief, fast healing or other improvement in symptoms or appearance and 23 reported inconclusive or negative outcomes. Direct application of light as a therapeutic intervention within the oral cavity (rather than photodynamic therapies, which utilize photosensitizing solutions) has thus far received minimal attention. Data from the limited studies that have been performed which relate to the oral cavity indicate that LLLT may be a reliable, safe and novel approach to treating a range of oral and dental disorders and in particular for those which there is an unmet clinical need. SIGNIFICANCE: The potential benefits of LLLT that have been demonstrated in many healthcare fields and include improved healing, reduced inflammation and pain control, which suggest considerable potential for its use in oral tissues.

Micronutrient modulation of NF-κB in oral keratinocytes exposed to periodontal bacteria.

Chronic periodontal diseases are characterised by a dysregulated and exaggerated inflammatory/immune response to plaque bacteria. We have demonstrated previously that oral keratinocytes up-regulate key molecular markers of inflammation, including NF-κB and cytokine signalling, when exposed to the periodontal bacteria Porphyromonas gingivalis and Fusobacterium nucleatum in vitro. The purpose of the current study was to investigate whether α-lipoic acid was able to abrogate bacterially-induced pro-inflammatory changes in the H400 oral epithelial cell line. Initial studies indicated that α-lipoic acid supplementation (1-4 mM) significantly reduced cell attachment; lower concentrations (<0.5 mM) enabled >85% cell adhesion at 24 h. While a pro-inflammatory response, demonstrable by NF-κB translocation, gene expression and protein production was evident in H400 cells following exposure to P. gingivalis and F. nucleatum, pre-incubation of cells with 0.5 mM α-lipoic acid modulated this response. α-Lipoic acid pre-treatment significantly decreased levels of bacterially-induced NF-κB activation and IL-8 protein production, and differentially modulated transcript levels for IL-8, IL-1ß, TNF-α and GM-CSF, TLR2, 4, 9, S100A8, S100A9, lysyl oxidase, NF-κB1, HMOX, and SOD2. Overall, the data indicate that α-lipoic acid exerts an anti-inflammatory effect on oral epithelial cells exposed to periodontal bacteria and thus may provide a novel adjunctive treatment for periodontal diseases.

Adjunctive daily supplementation with encapsulated fruit, vegetable and berry juice powder concentrates and clinical periodontal outcomes: a double-blind RCT.

AIM: A double-blind randomized controlled trial to determine whether dietary supplementation with fruit/vegetable/berry juice powder concentrates, simultaneously with non-surgical periodontal therapy, improved 2-month treatment outcomes. METHODS: Volunteers with chronic periodontitis were randomly assigned to one of three groups: fruit/vegetable (FV), fruit/vegetable/berry (FVB) or placebo. Supplements were taken daily during non-surgical debridement and maintenance and outcomes assessed at 2, 5 and 8 months after completion. Primary outcomes were mean probing pocket depth (PPD), clinical attachment gain, % sites bleeding on probing (% BOP) at 2 months. Adherence and plasma ß-carotene were determined. RESULTS: Sixty-one nutritionally replete (by serum biochemistry) volunteers enrolled and 60 (n = 20 per arm) completed the 2-month review. Clinical outcomes improved in all groups at 2 months, with additional improvement in PPD versus placebo for FV (p < 0.03). Gingival crevicular fluid volumes diminished more in supplement groups than placebo (FVB; p < 0.05) at 2 months, but not at later times. The % BOP (5 months) and cumulative plaque scores (8 months) were lowered more in the FV group (p < 0.05). CONCLUSIONS: Adjunctive juice powder concentrates appear to improve initial pocket depth reductions in nutritionally replete patients, where plasma micronutrient bioavailability is attainable. Definitive multicentre studies in untreated and treated patients are required to ascertain the clinical significance of such changes.